Impact of myeloid RIPK1 gene deletion on atherogenesis in ApoE-deficient mice

Background and aims: Targeting macrophage death is a promising strategy for stabilizing atherosclerotic plaques. Recently, necroptosis was identified as a form of regulated necrosis in atherosclerosis. Receptor-interacting serine/threonine-protein kinase (RIPK)1 is an upstream regulator of RIPK3, which is a crucial kinase for necroptosis induction. We aimed to investigate the impact of myeloid-specific RIPK1 gene deletion on atherogenesis. Methods: RIPK1F/FLysM-Cre+ApoE-/- and RIPK1+/+LysM-Cre+ApoE-/- mice were fed a western-type diet (WD) for 16 or 24 weeks to induce plaque formation. Results: After 16 weeks WD, plaque area and percentage necrosis in RIPK1F/FLysM-Cre+ApoE-/- mice were significantly decreased as compared to plaques of RIPK1+/+LysM-Cre+ApoE-/- mice. Moreover, plaques of RIPK1F/FLysM-Cre+ApoE-/- mice showed more apoptosis and a decreased macrophage content. After 24 weeks WD, plaque size and percentage necrosis were no longer different between the two groups. Free apoptotic cells strongly accumulated in plaques of RIPK1F/FLysM-Cre+ApoE-/- mice. In addition to apoptosis, necroptosis was upregulated in plaques of RIPK1F/FLysM-Cre+ApoE-/- mice. In vitro, TNF-α triggered apoptosis in RIPK1F/FLysM-Cre+ApoE-/-, but not in RIPK1+/+LysM-Cre+ApoE-/- macrophages. Moreover, RIPK1F/FLysM-Cre+ApoE-/- macrophages were not protected against RIPK3-dependent necroptosis. Conclusions: The impact of myeloid RIPK1 gene deletion depends on the stage of atherogenesis. At 16 weeks WD, myeloid RIPK1 gene deletion resulted in increased apoptosis, thereby slowing down plaque progression. However, despite decreased macrophage content, plaque and necrotic core size were no longer reduced after 24 weeks of WD, most likely due to the accumulation of free apoptotic and necroptotic cells.